Parathyroid hormone (PTH) is currently the only FDA-approved drug that replaces bone lost to osteoporosis. Despite its efficacy, there are serious limitations to its use because of high cost and concerns with treatment length1, 2. The quest for improved PTH efficacy has prompted the search for synergizers of PTH, i.e., pathways that accentuate PTH effects in the skeleton. One such synergistic stimulus is mechanical signaling. The molecular basis underlying the synergistic effects of PTH-induced anabolism and mechanically-induced anabolism are emerging3-6. One key factor that regulates both processes is the transcriptional repressor Nmp4 that dampens bone[unreadable]s response to PTH and to mechanical stimulation by inhibiting bone-anabolic genes7-14. Disabling Nmp4 in mice enhances PTH-mediated gains in bone and inhibits disuse-induced bone loss7, 12. Clinically, Nmp4 has been linked to osteoporosis susceptibility15, 16, indicating that changes in the function of this gene have consequences in the human population. Thus, inhibiting Nmp4 in patients may abbreviate and/or enhance teriparatide therapy and enhance the anabolic tonic derived from exercise, ultimately providing a prophylactic to disuse/postmenopausal osteoporosis. Recent data suggest that Nmp4 (1) suppresses recruitment to the osteoblast (OB) lineage, (2) represses OB differentiation, and (3) attenuates bone matrix synthesis7-14. In this competitive renewal application, we propose to investigate upstream mediators of Nmp4 activity and downstream targets of activated Nmp4, in the context of PTH/load synergism. In the cytoplasm Nmp4 associates with force-transducing adhesion proteins17, 18, therefore we hypothesize that Nmp4 activity is partly regulated by cell adhesion signaling. We propose Nmp4 suppresses the transcriptional capacity of two related PTH/load-activated pathways [unreadable] Bmp2/Smad and Wnt/&#56256;&#56321;-catenin. Furthermore, we hypothesize that activation of the R-Smads by both PTH and load synergistically enhances bone building by the Wnt/&#56256;&#56321;-catenin pathway and the novel adhesion receptor RAGE pathway;Nmp4 attenuates this synergy partly via suppression of &#56256;&#56321;-catenin and R-Smad activities10, 13. Specific Aim #1 is to determine whether Nmp4 deletion enhances bone sensitivity to PTH/load and improves PTH protection from disuse by subjecting genetically modified mice to PTH [unreadable] mechanical loading, or PTH [unreadable] mechanical disuse. Specific Aim #2 is to determine whether Nmp4-mediated suppression of &#56256;&#56321;-catenin and R-Smad activities is central to inhibiting PTH/loadgenerated anabolic signals;thus we will challenge osteoblasts from our engineered mice with (a) PTH and (b) oscillatory shear stress with or without hormone and monitor &#56256;&#56321;-catenin and R-Smad activities and anabolic gene expression. Specific Aim #3 is to determine how adhesion signaling mobilizes Nmp4 by altering osteoblast adhesion signaling and monitoring appropriate Nmp4 activation readouts. Our objective is to determine how Nmp4 regulates bone response to PTH/load synergism. The anticipated data will facilitate strategy development for enhancing PTH clinical utility.